Bottom Line:
Additionally, we found that BL-AD008-induced apoptosis was affected by the combination of AMPK and ZIPK.Then, we found that BL-AD008 bear its anti-tumor activities and induced apoptosis by targeting AMPK/ZIPK in vivo.In conclusion, these results demonstrate the ability of systems biology network to identify some key apoptotic kinase targets AMPK and ZIPK; thus providing a dual-target small molecule activator (BL-AD008) as a potential new apoptosis-modulating drug in future cervical cancer therapy.

ABSTRACTThe aim of this study was to discover a small molecule activator BL-AD008 targeting AMPK/ZIPK and inducing apoptosis in cervical cancer. In this study, we systematically constructed the global protein-protein interaction (PPI) network and predicted apoptosis-related protein connections by the Naïve Bayesian model. Then, we identified some classical apoptotic PPIs and other previously unrecognized PPIs between apoptotic kinases, such as AMPK and ZIPK. Subsequently, we screened a series of candidate compounds targeting AMPK/ZIPK, synthesized some compounds and eventually discovered a novel dual-target activator (BL-AD008). Moreover, we found BL-AD008 bear remarkable anti-proliferative activities toward cervical cancer cells and could induce apoptosis by death-receptor and mitochondrial pathways. Additionally, we found that BL-AD008-induced apoptosis was affected by the combination of AMPK and ZIPK. Then, we found that BL-AD008 bear its anti-tumor activities and induced apoptosis by targeting AMPK/ZIPK in vivo. In conclusion, these results demonstrate the ability of systems biology network to identify some key apoptotic kinase targets AMPK and ZIPK; thus providing a dual-target small molecule activator (BL-AD008) as a potential new apoptosis-modulating drug in future cervical cancer therapy.

Mentions:
To assess whether Fas-mediated pathway was activated in BL-AD008-treated HeLa cells, the levels of Fas, FasL, Fas-Associated protein with Death Domain (FADD) and caspase-8 were determined by Western blot analysis. The levels of Fas, FasL and FADD were markedly elevated and then there was obvious increase in the cleavage of caspase-8 after BL-AD008 administration (Figure 6A). Therefore, death receptor pathway is involved in BL-AD008-induced apoptosis. Next, we found that Bax expression was increased whereas Bcl-2 expression was decreased in HeLa cells. Moreover, we detected decrease of mitochondrial membrane potential by Rhodamin 123 staining in BL-AD008-treated HeLa cells (Figure 6B). It clearly indicates that BL-AD008-induced apoptosis in HeLa cells is mediated by a mitochondrial pathway. Then, we investigated the involvements of caspase-9 and caspase-3 in BL-AD008-induced apoptosis. Caspase-9 activation was determined by measurement of the active forms of caspase-9. The active form of caspase-3 was observed during BL-AD008 treatment (Figure 6B). These results suggest that mitochondrial pathway is also involved in BL-AD008-induced apoptosis. Moreover, we showed that the ratio of p-AMPK expression was increased in BL-AD008-treared HeLa cell apoptosis, and the ratio of ZIPK expression was also increased in this context. Thus, these results suggest that BL-AD008-induced apoptosis can be mainly affected by AMPK and ZIPK in HeLa cells (Figure 6C).

Mentions:
To assess whether Fas-mediated pathway was activated in BL-AD008-treated HeLa cells, the levels of Fas, FasL, Fas-Associated protein with Death Domain (FADD) and caspase-8 were determined by Western blot analysis. The levels of Fas, FasL and FADD were markedly elevated and then there was obvious increase in the cleavage of caspase-8 after BL-AD008 administration (Figure 6A). Therefore, death receptor pathway is involved in BL-AD008-induced apoptosis. Next, we found that Bax expression was increased whereas Bcl-2 expression was decreased in HeLa cells. Moreover, we detected decrease of mitochondrial membrane potential by Rhodamin 123 staining in BL-AD008-treated HeLa cells (Figure 6B). It clearly indicates that BL-AD008-induced apoptosis in HeLa cells is mediated by a mitochondrial pathway. Then, we investigated the involvements of caspase-9 and caspase-3 in BL-AD008-induced apoptosis. Caspase-9 activation was determined by measurement of the active forms of caspase-9. The active form of caspase-3 was observed during BL-AD008 treatment (Figure 6B). These results suggest that mitochondrial pathway is also involved in BL-AD008-induced apoptosis. Moreover, we showed that the ratio of p-AMPK expression was increased in BL-AD008-treared HeLa cell apoptosis, and the ratio of ZIPK expression was also increased in this context. Thus, these results suggest that BL-AD008-induced apoptosis can be mainly affected by AMPK and ZIPK in HeLa cells (Figure 6C).

Bottom Line:
Additionally, we found that BL-AD008-induced apoptosis was affected by the combination of AMPK and ZIPK.Then, we found that BL-AD008 bear its anti-tumor activities and induced apoptosis by targeting AMPK/ZIPK in vivo.In conclusion, these results demonstrate the ability of systems biology network to identify some key apoptotic kinase targets AMPK and ZIPK; thus providing a dual-target small molecule activator (BL-AD008) as a potential new apoptosis-modulating drug in future cervical cancer therapy.

ABSTRACTThe aim of this study was to discover a small molecule activator BL-AD008 targeting AMPK/ZIPK and inducing apoptosis in cervical cancer. In this study, we systematically constructed the global protein-protein interaction (PPI) network and predicted apoptosis-related protein connections by the Naïve Bayesian model. Then, we identified some classical apoptotic PPIs and other previously unrecognized PPIs between apoptotic kinases, such as AMPK and ZIPK. Subsequently, we screened a series of candidate compounds targeting AMPK/ZIPK, synthesized some compounds and eventually discovered a novel dual-target activator (BL-AD008). Moreover, we found BL-AD008 bear remarkable anti-proliferative activities toward cervical cancer cells and could induce apoptosis by death-receptor and mitochondrial pathways. Additionally, we found that BL-AD008-induced apoptosis was affected by the combination of AMPK and ZIPK. Then, we found that BL-AD008 bear its anti-tumor activities and induced apoptosis by targeting AMPK/ZIPK in vivo. In conclusion, these results demonstrate the ability of systems biology network to identify some key apoptotic kinase targets AMPK and ZIPK; thus providing a dual-target small molecule activator (BL-AD008) as a potential new apoptosis-modulating drug in future cervical cancer therapy.